Loss of Tolerance to Exogenous and Endogenous Factor VIII in a Mild Hemophilia A Patient With an Arg593 to Cys Mutation

A 42-year-old patient with mild hemophilia A developed spontaneous muscle hematomas 1 month after intense therapy with factor VIII concentrates. Factor VIII clotting activity was less than 1% and his factor VIII inhibitor was 10 Bethesda units (BU)/mL. The titer peaked at 128 BU despite daily infusions of factor VIII; 1 year later, the titer was 13 BU with no spontaneous bleeding for 4 months. The plasma inhibitor was 95% neutralized by factor VIII A2 domain but less than 15% neutralized by light-chain or C2 domain. His inhibitor did not cross-react with porcine factor VIII and was at least 10-fold less reactive to a series of hybrid factor VIII proteins in which human residues 484-508 are replaced by the homologous porcine sequence (Healey et al, J Biol Chem 270:14505, 1995). The inhibitor patient's DNA encoding his A2 domain and flanking sequences showed a C-T transition predicting Arg593 to Cys. Thirteen patients from 5 unrelated families with Cys593 have not developed inhibitors. Factor VIII clotting activity from one of them was inhibited similarly to diluted normal plasma by inhibitor patient plasma. In an homologous structure, ceruloplasmin (Zaitseva et al, J Biol Inorgan Chem 1:15,1996), the residue equivalent to Arg593, is in a loop distinct from residues 484-508. On solution phase immunoprecipitation with labeled factor VIII fragments, A2, light chain, and C2 domains bound. In contrast to typical immune responses to factor VIII in patients with severe hemophilia A, this patient's inhibitor was almost entirely reactive with common epitopes within the A2 domain whereas by more sensitive immunoprecipitation testing antibodies to light chain epitopes were also present. Accordingly, immune responsiveness to exogenous factor VIII (antigen burden) appears to be more critical than his endogenous, hemophilic factor VIII to his developing high-titer anti–factor VIII antibodies and loss of tolerance to both native and hemophilic factor VIII proteins.